This application describes a broad program designed to study the mechanism of action of folate antagonists and folate biochemistry and pharmacology from a basic level with extension into the clinical level. Studies of dihydrofolate reductase from several human tissues, both normal and neoplastic, will be carried out. Comparative structural, physical and kinetic studies are planned to examine potential differences between these enzyme sources. Regulation of certain folate enzymes, in particular, dihydrofolate reductase, methionine synthetase and methylene tetrahydrofolate reductase will be studied. Folate coenzyme function and metabolism, in particular folate polyglutamates, and N10 formy folic acid will also be investigated. New folate antagonists, in particular "non-classical" antifolates, and a folate cleaving enzyme, carboxypeptidase G1 are also being studied both in the laboratory and in early trials in patients with malignant disease. The goal of these studies remains the improvement of therapy of neoplastic disease in man; as promising approaches are developed, they are extended to the clinical level and appropriately monitored.